The present invention relates to a micro-chemistry reaction method which mainly manipulates a droplet including particles responding to a magnetic force such as a magnetic body or particles including the above-mentioned particles by applying a magnetic field to the above-mentioned particles on a solid-phase surface or near the solid-phase surface; and execute operations such as preparative isolation, displacement, mixture, heating, and cooling of the fluid necessary for a chemical reaction.
For an efficient control and analysis of a reaction of the chemical reaction system and a minimization of the chemical reaction system, especially, a biochemical reaction system, a micro-chemical reaction system, i.e. μ-TAS (Micro Total Analysis System) structure which can be used for the screening of medical diagnostics or the development of a new medicine, an identification test in a forensic medicine field, environmental monitoring and so on, is expected. Recently, a μ-TAS device has been studied by developing a micro flow channel or valve on a silicon or glass substrate by a microfabrication technology in a semiconductor manufacturing field.
Characteristics required for the chemical reaction on a microchip such as the μ-TAS device are efficiency of the reaction and shortening of the analysis time due to the minimization; high throughput processing due to the integration; reduction of burdens on an examinee due to the reduction of the collection quantity of a specimen material; improvement of analytical precision due to automation; reduction of environmental burdens due to the control of the amount of reagent or waste fluid; and reduction of cost. On the other hand, in addition to easiness of chip manufacturing, especially, mass production; low manufacturing cost; and handling ability, reducing size and weight of the equipment for conducting the reaction and analysis are required in order to realize the device. Furthermore, low cost and higher reliability are required for the device.
A conventional μ-TAS chip is provided with the micro flow channel on the silicon or glass substrate using semiconductor manufacturing technology, and can obtain high processing accuracy and various measures of the fluid control in a micro area. However, the conventional μ-TAS chip is impractical since a peripheral device, which conducts the reaction, tends to become large-scale and complicated in addition to a difficulty to control the cost for expensive materials or high-accuracy processing. Especially, in a genetic diagnosis in a medical field which can not cause cross-contamination, the size and weight of analyzing equipment are required to be small, and the chip has to be a disposable μ-TAS chip which can conduct the reaction in a sealed atmosphere.
On the other hand, because of the expectation of the above-mentioned application, in order to reduce the manufacturing cost and be disposable, a μ-TAS chip using a resin substrate has been studied. However, when a conventional μ-TAS approach is adopted, the establishment and maintenance of the processing accuracy when a micro flow channel, micropump, valve, mixer and so on are formed by the resin substrate, are difficult to achieve, compared to the silicon or glass substrate. However, a resin chip has an excellent applicability to a bio-genic substance such as nucleic acid and protein, and a biological sample such as a body tissue or cell in addition to that raw materials thereof are affordable, so that the resin chip is very advantageous as a material for the disposable μ-TAS chip.
A technology for transporting a minute amount of liquid in a μ-TAS field has been numerously reported. As a driving method with respect to the solution sending the minute amount of liquid, a method in which liquid or gas from a drive source of a device external portion is a driving medium by a syringe-type pump; or an electrical method using an electrostatic force or dielectrophoresis have been reported.
When utilization for the biochemical reaction, especially, diagnosis in clinical medical care is considered, in many cases, the amount of the sample at the starting time of the reaction is minute. Also, in order to reduce the burdens of the examinee, the reduction of the collection quantity of the specimen materials will be furthermore required in the future, so that a micro-amount biochemical reaction system using a μ-TAS technology is required. Also, when a large-sized screening is considered, high throughput processing of the system due to integration and parallelization is required. In the conventional minimization of the biochemical reaction, the micro flow channel was built on a silicon or glass substrate, and the solution sending, distribution, mixture and the like were conducted. However, cost and labor hours for providing the microfabrication on the substrate and also pasting substrates are required, and even if a reaction chip becomes smaller, a high-accuracy operation due to an external solution sending equipment of the solution sending drive source is required, so that the overall system becomes large-scale and impractical.
On the other hand, when the reaction chip is made disposable, the development of a resinous chip is essential; however, in order to directly mold a microflow system which is designed by materials except for resin such as a silicon substrate and the like on the resin chip, it is difficult to ensure a processing accuracy of a micrometer-order, and it is required to be designed while considering the shrinkage and transformation when molded. Accordingly, there are many problems in building the micro flow channel, micro solution sending pump, and micro valve on the resin substrate, and there are also many problems in promoting mass production.
For transporting the minute amount of liquid in a droplet state, a method electrically transporting the droplet which is formed when, for example, a minute amount of oil is added to water or a minute amount of water is added to oil, has been reported. An electrostatic carrier (refer to a Japanese non-patent document 1: “International Symposium on Microchemistry and Microsystems 2001” by Tomohiro Taniguchi, et al., 2001, pp. 104-105) using an electrostatic force two-dimensionally operates various droplets, and conducts the micro chemical reaction. However, it is required to arrange a micro electrode and also provide a high-accuracy insulation processing in the device manufacturing in addition to requiring high voltage. Accordingly, it is difficult to mass produce, and costs for a disposable device are not affordable.
On the other hand, a chemical reaction operation of the droplet including the magnetic body has also been reported (refer to Japanese non-patent document 2: “Sensors and Actuators B” by Mitsuhiro Shikida, et al., 2006, Vol. 113, pp. 563-569, and patent document 3: International Patent Publication No. 05/069045). In the above-mentioned documents, the following method has been proposed. The sample material or a material connected with the sample material is connected on the surfaces of magnetic body particles beforehand, and the magnetic body particles are displaced by a magnet and united with a droplet (for example, enzyme liquid and the like) which includes the other reagent, so that a chemical reaction is conducted. However, an object of this method is promoting the reaction process of the material connected on the surface of the magnetic body by transporting the magnetic body itself. Also, a fluid control structure such as a dividing wall is provided on a glass device, and the μ-TAS chip on the resin substrate which does not require a special processing which is a problem to be resolved by this invention, has not been obtained. Therefore, the method is directed to one method of the conventional μ-TAS device, and differs from μ-TAS technology with a novel concept wherein the micro flow channel and micropump are not required, which is the intention of this invention.
As a microchip using the magnetic body particles, there is a chip extracting and purifying the nucleic acid from a sample on a substrate, and conducting a gene amplification reaction (refer to the Japanese patent document 1: Japanese Patent Publication (TOKKAI) No. 2006-61031). In this chip, the sample and magnetic body particles are filled into a refined well; the nucleic acid in the sample is attached to the magnetic body particles; and the nucleic acid is purified by displacing the magnetic body to the other refined well using fluctuations of the magnetic field. The magnetic body wherein the nucleic acid is purified is displaced to a nucleic-acid amplification well on the chip; and the magnetic body displaced to the nucleic-acid amplification well is heated by induction heating, so that the nucleic acid attached to the magnetic body is amplified. In this chip, a reaction field such as purification and nucleic-acid amplification is within the well provided in the substrate, and in order to process the above-mentioned well on the substrate, high-accuracy and complicated microfabrication is required. Even in the above-mentioned reference examples, substrate processing is required.
In addition, the reference examples have a structure specialized in a standard protocol of the gene amplification reaction, so that they cannot respond flexibly to various biochemical reaction protocols. As in the case of the Japanese non-patent document 2, this chip is based on the assumption that the example material (DNA) is attached to the surface of the magnetic body, and this is also one method of the conventional μ-TAS device. Accordingly, the chip differs from the μ-TAS technology with the novel concept wherein the micro flow channel and micropump are not required, which is the intension of this invention.
Also, there is a method of conducting a PCR (polymerase chain reaction) within the droplet by repeatedly displacing and placing the droplet formed from PCR reagent by the electrostatic transportation among multiple heated areas (refer to Japanese patent document 2: International Patent Publication No. 03/067875).
The present invention provides a method and device which do not require to place or construct a fluid control element such as a pump, valve, mixer and the like inside a reaction receptacle, which was required for a conventional μ-TAS, so that chemical reaction operations such as the solution sending, preparative isolation, mixture, dilution, agitation and temperature control of liquid can be conducted, and a possible chemical reaction after the above-mentioned operations can be conducted.
Further objects and advantages of the invention will be apparent from the following description of the invention.